Dry eye affects over six million people in the United States alone and may be classified as aqueous production-deficient (AD) and evaporative, which is associated with meibomian gland dysfunction (MGD). Both mechanisms of development of dry eye share the common feature of instability of the tear film (TF) with rapid tear break-up time, suggesting there may be a shared structural abnormality of the TF that is responsible for the instability. Our spectroscopic studies have identified several potential abnormalities in the TF lipid composition that may contribute to TF instability including terpenoids, saturation, protein and cholesteryl esters. We also identified hydrocarbon conformational abnormalities with dry eye. For example, when terpenoid levels in meibum are low as in MGD, the TF is unstable and patients have the signs and symptoms of dry eye. When terpenoids are restored with azithromycin treatment, TF stability is restored and patients no longer have the signs and symptoms of dry eye. We have the best chance of testing which moieties are important to TF stability by studying samples with a wide range of TF stabilities. We propose to study samples from patients with dry eye (both AD and MGD) and samples from Asians because they have an 80% higher incidence of dry eye compared to Caucasians. We will also compare compositional changes in meibum and TF lipid when the signs and symptoms of dry eye are ameliorated with treatment. TF lipid and meibum will be studied since their composition is slightly if not significantly different. We propose to use spectroscopy, Raman, IR and NMR, to determine how TF composition relates to tear film stability measured in vitro and in vivo on the same sample. The TF stability parameters we propose to measure in vitro include: surface pressure-area isotherms, isocycles, surface rheology, surface Raman and quantitative Brewster angle microscopy to assess the meibum's lateral elasticity and capability of compressing and spreading during dynamic area changes. We propose to measure in vivo several parameters including tear osmolarity, break-up time and blink rate. Natural and experimental differences in TF lipid and meibum amount and composition (especially terpenoids, saturation, protein and levels of cholesteryl esters), hydrocarbon conformation, and lipid-protein interactions, will be correlated to TF stability using principle component analysis t determine if and which moieties in meibum are important to TF stability, with the ultimate goal of developing therapies to enhance TF stability.